This proposed project focuses on quantifying the spatial and temporal diversity in cortical oxygen metabolism and neurovascular coupling, and informing next generation of biophysical models of the Blood Oxygen Level Dependent (BOLD) functional Magnetic Resonance Imaging (fMRI) measurements by integrating new technological and conceptual approaches. Broad and long term objectives of this application are (1) to bridge understanding of brain in action at multiple scales in spatial (subcapillary to whole- brain) and temporal (s to months) domains, (2) to contribute discovering diversity of hemodynamic responses across cortical tissue layers, functional areas, and microvascular branches, and (3) to support the modelling efforts of BOLD signal by to quantifying hemodynamic responses in cortex of healthy-aging awake mice. Realizing these objectives would contribute to the joined efforts towards improving human health through accomplishing BRAIN Initiative goals of understanding the brain in action, discovering diversity, mapping/linking activity across spatial and temporal scales, and understanding the biophysical basis of fMRI signal. The specific aims of the proposed project can be summarized as: (1) to develop faster, deeper, longitudinal measurements of the cerebral blood flow (CBF), partial pressure of oxygen (pO2), and cerebral metabolic rate of oxygen (CMRO2), (2) to quantify the diversity of CMRO2 and CBF responses to functional activation across different cortical functional areas, cortical layers, and microvasculature types in healthy-aging mouse, and (3) to quantify the effects of standard fMRI calibrations (e.g. hypercapnia, hyperoxia, and caffeine) on the CMRO2 and CBF at rest and during functional activation. The research design and methods that will be used in this project will include (1) developing faster and deeper chronic imaging of absolute oxygen concentration to assess relation between cortical pO2, CMRO2, and blood flow changes during brain activation across the cortical areas and layers, and (2) realistic measurements of the oxygenation, blood flow, and metabolism responses to functional activation at the microvascular scales in healthy-aging mice and in response to calibration procedures that are commonly used in BOLD fMRI imaging. The research environment of Dr. Sencan (the Athinoula A. Martinos Center for Biomedical Imaging at Massachusetts General Hospital and Radiology Department in Harvard Medical School) is equipped with state-of-the-art technologies required to execute the proposed project and is rich with opportunities for training in neuroscience, career development, and interdisciplinary collaborations. She has a team of mentors, collaborators and advisors with diverse and strong expertise, who are committed to support Dr. Sencan?s research, and her career development. All these factors will facilitate the successful execution of the proposed project, the completion of Dr. Sencan?s training in neuroimaging, and her transition to an independent tenure-track faculty position.